The invention relates to a new process for the production of perbenzylated 1-O-glycosides of general formula I, which is characterized in more detail in the claims. The process according to the invention starts from economical starting materials, provides good yields and allows the production of perbenzylated saccharides with 1-O-functionalized side chains on an enlarged scale.
Perbenzylated saccharide derivatives are valuable intermediate products in synthetic chemistry. Primarily pharmaceutical chemistry uses such components very frequently, since many highly potent and selective pharmaceutical agents carry sugar radicals. Thus, for example, in the Journal of Drug Targeting 1995, Vol. 3, pp. 111-127, applications of the so-called xe2x80x9cglycotargetingxe2x80x9d are described. So-called xe2x80x9cmulti-antennary sugar chainsxe2x80x9d are described in Chemistry Letters 1998, p. 823. By clustering sugar units, the carbohydrate-receptor-interaction in the case of the cell-cell-interaction is considerably improved. The synthesis of galactosides with high affinity to the asialoglycoprotein receptor is published in J. Med. Chem. 1995, 38, p. 1538 (see also Int. J. Peptide Protein Res. 43, 1994, p. 477). Here, derivatized galactoses with functionalized side chains are produced, which then can be suspended on various other molecules. A good survey on the use of saccharides as a basis of glycobiology has been provided in Acc. Chem. Res. 1995, 321. Also, for the synthesis of LewisX mimetic agents (Tet. Lett. Vol. 31, 5503), functionalized monosaccharides are used as precursors (see also JACS 1996, 118, 6826).
The use of derivatized monosaccharides as intermediate stages for potential pharmaceutical agents has been well represented in Current Medicinal Chemistry, 1995, 1, 392. Perbenzylated-1-OH-sugar derivatives (galactose, glucose) are also used in the synthesis of heart-active glycosides (digitoxin-conjugates). The 1-O-glycosylation is carried out here via trichloroacetimidate and BF3-catalysis (J. Med. Chem. 1986, 29, p. 1945). For the production of immobilized sugar ligands (e.g., linkage to HSA), functionalized, protected monosaccharides are used (Chemical Society Reviews 1995, p. 413).
It is the purpose of a group of syntheses to introduce additional functionality into a sugar molecule via a 1-O-glycosylation reaction. Here, primarily terminal COOHxe2x80x94, amino- or OHxe2x80x94 groups are of interest, since the latter can be further reacted in subsequent steps.
The production of 1-O-glycosides is carried out in most cases according to standard methods, such as, e.g., according to the trichloroacetimidate methods described by Koenigs-Knorr, Helferich or by R. R. Schmidt [W. Koenigs and E. Knorr, Ber. dtsch. Chem. Ges. 34 (1901) 957; B. Helferich and J. Goendeler, Ber. dtsch., Chem. Ges. 73, (1940) 532; B. Helferich, W. Piel and F. Eckstein, Chem. Ber. 94 (1961), 491; B. Helferich and W. M. Mxc3xcller, Chem. Ber. 1970, 103, 3350; G. Wulff, G. Rxc3x6hle and W. Krxc3xcger, Ang. Chem. Internat. Edn., 1970, 9, 455; J. M. Berry and G. G. S. Duthon, Canad. J. Chem. 1972, 50, 1424; R. R. Schmidt, Angew. Chem. 1986, 98, 213.]
A feature that is common to all of these methods is that the 1-hydroxyl group is converted into a reactive form that is ultimately used as a leaving group. Under Lewis acid catalysis (partially in stoichiometric amount), the actual reaction is carried out with an alcohol to 1-O-glycoside. For such reactions, numerous examples are provided in the literature.
In the production of immunostimulant KRN-7000 (Kirin Brewery), the condensation of tetra-O-benzyl-xcex2-D-galactopyranosyl-bromide with a primary alcohol, whose hydroxyl group sits at the end of a di-hydroxy-amido-C-chain (in DMF/toluene under Lewis acid catalysis), is thus a central step (Drug of the Future 1997, 22(2), p. 185). In Japanese Patent JP 95-51764, the reaction of 1-O-acetyl-2,3,4-tri-O-benzyl-L-fucopyranose with polyoxyethylene-30-phytosterol (BPS-30, NIKKO Chem., Japan) under trimethyl-silylbromide/zinc triflate catalysis was described. In Bull. Chem. Soc. 1982, 55(4), pp. 1092-6, 1-O-glycosylations of perbenzyl-sugars under titanium tetrachloride catalysis in dichloromethane are described.
In Liebigs Ann. Org. Bioorg. Chem.; En; 9; 1995; 1673-1680, the production of 3,4,5-trisbenzyloxy-2-benzyloxymethyl-6-(2-hexadecyloxyethoxy)-tetrahydropyran is described. Starting from 2,3,4,6-tetra-O-benzyl-D-glucopyranose, the 1-O-glycosylation is performed with use of Bu4NBr, CoBr2, Me3SiBr and a molecular sieve in methylene chloride within 60 hours.
A tetrabenzyl derivative, which contains a terminal carboxyl group that is protected as a methyl ester, is described in Carbohydr. Res.; EN; 230; 1; 1992; 117. The carboxyl group can then be released and further reacted. For glycosylation, silver carbonate is used in dichloromethane. The use of expensive silver carbonate limits the batch size and makes an economical up-scaling almost impossible. The same problem applies for the compound below, which was described in Tetrahedron Lett. 30, 44, 1989, p. 6019. Here, 2,3,4,6-tetra-O-benzyl-D-mannosyl-bromide in nitromethane is reacted with 2-benzyloxyethanol with the aid of mercury cyanide to form 1-O-glycoside. The use of mercury cyanide in pilot-plant installations is problematical in nature and can be rejected from the environmental-political standpoint.
The substance libraries for the high-capacity-screening described most recently very frequently use saccharides (Angew. Chemie 1995, 107, 2912). Here, the purpose is to have sugar components present in protected form, which carry a functional group, such as, e.g., xe2x80x94COOH, or xe2x80x94NH2, which can be reacted in, e.g., an automated synthesis. The components, which are used in this respect, were described by Lockhoff, Angew. Chem. 1998, 110 (24), p. 3634. Primarily the 1-O-acetic acid of perbenzyl-glucose is important here. The production is carried out over two stages, via trichloroacetimidate and reaction with hydroxyacetic acid ethyl ester, BF3 catalysis in THF and subsequent saponification with NaOH in MeOH/THF. The total yield over two stages is only 59%, however.
The 1-O-ethyl acetate that has been intermediately processed in this case is obtained in EP 882733 by reaction of 2,3,4,6-tetra-O-benzylglucose with hydroxyacetic acid ethyl ester in the presence of catalytic amounts of p-toluenesulfonic acid by reflux-boiling in benzene, but without data on the yield.
In the same publication, the production of a 1-O-(aminoethyl)-glycoside of the perbenzylated glucose is also described. The reaction is carried out, also starting from trichloroacetimidate, by reaction with N-formylaminoethanol under BF3-catalysis in THF and subsequent saponification in MeOH/THF. The total yield is also relatively low here; it is 45%.
A 1-O -(aminoethyl) derivative of perbenzylxylose passes through as an intermediate product in Carbohydrate Research 1997, 298, p. 173. The synthesis is very lengthy, however, since it starts from 1-bromo-peracetate of xylose. The actual 1-O-glycosylation is carried out via a 1-phenylthioether, which is reacted with 2-azidoethanol under DMTST catalysis (=dimethyl (methylthio)-sulfonium-triflate) in dichloromethane (total number of stages: 7). The total yield is not suitable for an industrial application with less than 40%.
In the survey article by R. R. Schmidt in Angew. Chem. 1986, 98, pp. 213-236, direct reactions of 1-OH-perbenzyl-glucose and -ribose with 2-haloesters and triflates are described. As a base, sodium hydride in THF or benzene is used (Chem. Ber. 1982, 115); the yields are between 40 and 55%. The use of sodium hydride in dioxane or potassium-tert-butylate in THF (both at room temperature) is also described for 1-O-alkylation with triflates (Angew. Chem. 1986, 98, p. 218). The anhydrous reaction conditions that are to be followed most strictly represent a large hurdle in up-scaling such alkylations.
All processes known to date have the great disadvantage that an up-scaling of the process cannot be achieved easily. The use of Lewis acids in 1-O-glycosylation and sodium hydride in 1-O-alkylation already requires anhydrous reaction conditions, which in large batches is always associated with difficulties. The working-up and disposal of reaction adjuvants (Hg/cyanide/etc.) is also a problem in many cases.
The object of the invention was therefore to provide a process with which perbenzylated saccharides with 1-O-functionalized side chains can be produced at a reasonable price and in an ecologically beneficial way on an enlarged scale.
The object of the invention is achieved according to the process indicated in the claims, with which perbenzylated 1-O-glycosides of general formula I 
can be produced. According to the definition of the invention, sugar1 in general formula I means a monosaccharide functionalized in 1-OH position, whereby in this connection, it can also be a deoxy sugar, which contains an H-atom instead of one or more OH groups. In a preferred embodiment of the invention, the sugar in general formula I means a monosaccharide with 5 or 6 C-atoms, e.g., glucose, mannose, galactose, ribose, arabinose or xylose or deoxy sugars thereof, such as, for example, 6-deoxygalactose (fucose) or 6-deoxy-mannose (rhamnose).
Radical R represents the benzyl group that is present in at least two places based on the monosaccharide that is used or its deoxy form, and is present accordingly in several places with use of di-, tri- or polysaccharides.
Radical X means xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94COOxe2x80x94 or xe2x80x94NHxe2x80x94. In the results of the process according to the invention, alcohols, carboxylic acids, or amines of general formula I are thus obtained.
Radical L can mean a straight-chain, branched, saturated, or unsaturated C1-C30 carbon chain, which optionally is interrupted by 1-10 oxygen atoms, 1-3 sulfur atoms; 1-2 phenylene groups, 1-2 phenylenoxy groups, 1-2 phenylenedioxy groups; a thiophene radical, pyrimidine radical or pyridine radical; and/or optionally is substituted with 1-3 phenyl groups, 1-3 carboxy groups, 1-5 hydroxy groups, 1-5 Oxe2x80x94C1-C7 alkyl groups, or 1-3 amino groups; 1-3 CF3 groups, or 1-10 fluorine atoms. In terms of the invention, preferred radicals L are 
whereby xcex3 means the interface site to the sugar, and xcex4 is the interface site to radical X. An especially preferred linker L is the xe2x80x94CH2 group.
For the production of perbenzylated 1-O-glycosides of general formula I, a perbenzylated 1-OH sugar of general formula II 
in which sugar, R and n have the above-indicated meaning, is dissolved in a water-immiscible organic solvent and reacted with an alkylating reagent of general formula III
Nuxe2x80x94Lxe2x80x94X Sg xe2x80x83xe2x80x83(III), 
in which Nu means a nucleofuge, L and X have the above-mentioned meaning, and Sg is a protective group, in the presence of a base and optionally a phase transfer catalyst. As a nucleofuge, for example, the radicals xe2x80x94Cl, xe2x80x94Br, xe2x80x94I, xe2x80x94OTs, xe2x80x94OMs, xe2x80x94OSO2CF3, xe2x80x94OSO2C4F9 or xe2x80x94OSO2C8F17 can be contained in the alkylating reagent of general formula III.
Protective group Sg is a common acid- or amine-, hydroxy- or thiol protective group, depending on whether X means the radical xe2x80x94Oxe2x80x94, xe2x80x94COOxe2x80x94 or xe2x80x94NHxe2x80x94. These protective groups are well known to one skilled in the art (Protective Groups in Organic Syntheses, Second Edition, T. W. Greene and P. G. M. Wuts, John Wiley and Sons, Inc., New York 1991).
The reaction according to the invention can be carried out at temperatures of 0-50xc2x0 C., preferably 0xc2x0 C. to room temperature. The reaction times are 10 minutes to 24 hours, preferably 20 minutes to 12 hours.
The base is added either in solid form, preferably fine-powder, or as 10-70%, preferably 30-50%, aqueous solution. NaOH and KOH are used as preferred bases.
As organic, water-immiscible solvents, for example, toluene, benzene, CF3-benzene, hexane, cyclohexane, diethylether, tetrahydrofuran, dichloromethane, MTB or mixtures thereof can be used in the alkylating process according to the invention.
As phase transfer catalysts, the quaternary ammonium or phosphonium salts that are known for this purpose or else crown ethers, such as, e.g., [15]-crown 5 or [18]-crown 6, are used in the process according to the invention. Quaternary ammonium salts with four hydrocarbon groups that are the same or different on the cation, selected from methyl, ethyl, propyl, isopropyl, butyl or isobutyl, are preferably suitable. The hydrocarbon groups on the cation must be large enough to ensure good solubility of the alkylating reagent in the organic solvent. According to the invention, N(butyl)4+-Clxe2x88x92, or N(butyl)4+-HSO4xe2x88x92, but also N(methyl)4xe2x80x2-Clxe2x88x92 is especially preferably used.
After the reaction is completed, the working-up of the reaction mixture can be carried out by isolation of the still protected end product and subsequent usual cleavage of the protective group to the end product of general formula I. It is preferred, however, not to isolate the still protected end product but rather to remove the solvent, to take up the residue in a new solvent that is suitable for the cleavage of the protective group and to perform the cleavage here. The procedure for cleavage of the protective group and for regeneration of the acid, amino, hydroxy or thiol group is well known to one skilled in the art.
In protective group Sg, e.g., this is an acid protective group, which blocks the acid proton of the carboxy group, thus, e.g., methyl, ethyl, benzyl or tert-butyl, such that the acid is usually regenerated by alkaline hydrolysis. In the process of the invention, for this case after the solvent is removed from the alkylating reaction, the residue is now taken up in a new solvent, e.g., methanol, ethanol, tetrahydrofuran, isopropanol, butanol or dioxane. An aqueous solution is then added to a base, and the alkaline hydrolysis is performed at temperatures of 0-100xc2x0 C.
As hydroxy protective groups, e.g., benzyl, 4-methoxybenzyl, 4-nitrobenzyl, trityl, diphenylmethyl, trimethylsilyl, dimethyl-tert-butylsilyl, or diphenyl-tert-butylsilyl groups are suitable.
The hydroxy groups can also be present, e.g., as THP-ethers, xcex1-alkoxyethylethers, MEM-ethers or as esters with aromatic or aliphatic carboxylic acids, such as, e.g., acetic acid or benzoic acid. In the case of polyols, the hydroxy groups can also be protected in the form of ketals with, e.g., acetone, acetaldehyde, cyclohexanone or benzaldehyde.
The hydroxy protective groups can be released according to the literature methods that are known to one skilled in the art, e.g., by hydrogenolysis, acid treatment of ethers and ketals, alkali treatment of esters or treatment of silyl protective groups with fluoride (see, e.g., Protective Groups in Organic Syntheses, Second Edition, T. W. Greene and P. G. M. Wuts, John Wiley and Sons, Inc., New York, 1991).
The thiol groups can be protected as benzyl-ethers, which can be cleaved with sodium in ammonia or boiling ethanol (W. J. Patterson, v. du Vigneaud, J. Biol. Chem. 111:393, 1993). S-tert-Butyl ethers can be easily cleaved with hydrogen fluoride/anisole at room temperature [S. Salzakibona et al., Bull. Chem. Soc. Japn, 40:2164, (1967)]. S-Benzyloxycarbonyl derivatives can be easily cleaved by concentrated ammonia solution at room temperature (A. Berger et al., J. Am. Chem. Soc., 78:4483, 1956). S-Benzyloxycarbonyl derivatives of trifluoroacetic acid are cleaved only at boiling point [L. Zervas et al., J. Am. Chem. Soc., 85:1337 (1963)].
The NH2 groups can be protected and released again in a variety of ways. The N-trifluoroacetyl derivative is cleaved by potassium or sodium carbonate in water [H. Newman, J. Org. Chem., 30:287 (1965), M. A. Schwartz et al., J. Am. Chem. Soc., 95 G12 (1973)] or simply by ammonia solution [M. Imazama and F. Eckstein, J. Org. Chem., 44:2039 (1979)]. The tert-butyloxycarbonyl derivative is equally easy to cleave: stirring with trifluoroacetic acid suffices [B. F. Lundt et al., J. Org. Chem., 43:2285 (1978)]. The group of NH2-protective groups to be cleaved hydrogenolytically or reductively is very large: The N-benzyl group can be cleaved easily with hydrogen/Pdxe2x80x94C [W. H. Hartung and R. Simonoff, Org. Reactions VII, 263 (1953)], which also applies for the trityl group [L. Zervas, et al., J. Am. Chem. Soc., 78:1359 (1956)] and the benzyloxycarbonyl group [M. Bergmann and L. Zervas Ber. 65:1192 (1932)].
Of the silyl derivatives, the easily cleavable tert-butyldiphenylsilyl compounds [L. E. Overman et al., Tetrahedron Lett., 27:4391 (1986)] and the 2-(trimethylsilyl)-ethyl carbamate [L. Grehn et al., Angew. Chem. Int. Ed. Engl., 23:296 (1983)] and the 2-trimethylsilylethanesulfonamide [R. S. Garigipati and S. M. Weinreb, J. Org. Chem., 53:4134 (1988)] are used, which can be cleaved with fluoride ions. Especially easily cleavable is the 9-fluorenylmethyl-carbamate: The cleavage is carried out with amines such as piperidine, morpholine, 4-dimethylaminopyridine, but also with tetrabutylammonium fluoride [L. A. Corpino et al., J. Org. Chem., S5:1673 (1990); M. Ueki and M. Amemiya, Tetrahedron Lett., 28:6617 (1987)].
The isolation of the end product of general formula I (alcohol, thiol, amine or carboxylic acid) that is obtained is also carried out according to common methods that are well known to one skilled in the art.
Thus, for example, in the case of the acid protective group, the solvent is evaporated from the hydrolysis reaction, and the residue is taken up in an aprotic solvent.
By acidification with an aqueous acid solution, the pH is set at about 2-4, and then the organic phase is separated. Using crystallization or chromatography, the perbenzylated 1-O-glycoside can now be obtained.
The compounds of general formula I that are obtained optionally also can be converted into their salts in the usual way.
The yields of the compounds of general formula I, which can be achieved with the process according to the invention, are good. For known compounds in which a comparison with the prior art is possible, they exceed the yields of the prior art. Thus, for example, for 1-O-acetic acid of perbenzylated glucose, a total yield of 59% is described in Angew. Chem. 1998, 110 (24), p. 3634 with the process that is mentioned there, while according to the invention, the yield for this compound is 82% over two stages (cf. Example 7 of this application). The production of the compound of Example 12 of this application is also described in this publication. While the yield of this compound according to the invention is 78% over 2 stages, only 45% is achieved with the process that is described in the publication.
In addition to the high yields, the process according to the invention also offers the advantage that it starts from economical starting materials, makes possible a scale-up of the process, and allows an easy isolation of the end products.
The starting materials are commercially available products or can be obtained easily from commercially available precursors. Tetra-2,3,4,6-O-benzyl-D-glucopyranose thus can be obtained in the case of Fluka AG, Duchs, Switzerland. In the case of Fluka, methyl-D-manno-pyranoside and methyl-D-galactopyranoside are also catalog items. By benzylation and cleavage of the glycoside, 2,3,4,6-tetra-O-benzyl-D-mannose or -galactose can be obtained.
The perbenzyl-1-OH derivatives of the pentoses (ribose, arabinose), hexoses and deoxyhexoses (rhamnose, fucose) can be obtained via the sequence of methylglycoside-perbenzyl-methylglycoside-perbenzyl-1-OH-saccharide.
The compounds that are produced according to the invention are valuable intermediate products in synthetic chemistry. They can thus be used, for example, in the synthesis of carbohydrate dendrimers, for synthesis of NMR contrast media and for introducing sugar radicals into pharmaceutical agents.
The process according to the invention is to be explained in more detail below in the embodiments.